Thioredoxin, one class of protein thiol-disulfide oxidoreductase, was first discovered in Escherichia coli in 1964 as a coenzyme of ribonucleotide reductase, which is essential for DNA synthesis. Thioredoxins are commonly found in nearly all biological species. It is known that the thioredoxin, together with other enzymes such as peroxyredoxin serving as an electron acceptor, functions as a key anti-oxidase in organisms to show enable various physiological activities, such as control of gene expression, control of cell proliferation, control of cell death, elimination of reactive oxygen species in the cell, inhibition of neutrophil activation, and inhibition of neutrophil migration. The role of thioredoxin as an important protective agent against oxidative stress has also been elucidated through its resistance to symptoms and diseases closely associated with oxidative stress, such as aging, ischemic disorder, acute lung failure, and diabetes mellitus. Because of these and other useful physiological activities, thioredoxins are considered to be useful in applications such as functional food and medicaments.
The use of thioredoxin in food and medicaments requires a high level of safety because it involves ingestion by or administration to humans. In this connection, the use of yeast, which has long been used in the human diet through applications such as bread and beer production, is thought to be desirable for the production of thioredoxin intended for food and medicaments. However, because the thioredoxin biosynthesized in yeast accumulates in the yeast cell, thioredoxin has been conventionally obtained from yeast via the process of disrupting the cultured yeast cell, using physical means such as glass beads or a French press, or using a cell-wall-digesting enzyme. A problem of conventional thioredoxin production processes, then, is that the disruption of yeast cells introduces a large amount of contaminant proteins other than thioredoxin, with the result that the purity of the product thioredoxin is inevitably low. As a process for producing high-purity thioredoxin using yeast, a method in which predetermined membrane fractionation is performed after disrupting the yeast cells has been proposed. However, this method still falls short in terms of thioredoxin purity. The purity of the thioredoxin obtained can be improved by combining various types of conventional purification techniques such as chromatography, salting out, and membrane fractionation. However, considering the large numbers of cells required as a starting material in actual production, it is not practical to improve the purity of thioredoxin solely using these purification techniques.
As described above, the technique of producing high purity thioredoxin, as it currently stands, is not sufficient to provide an industrially feasible process.
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